Measles virus (MV) infection causes an acute childhood disease and, in some cases, can lead to severe neurological sequelae and death. Specific antiviral therapy for measles is urgently needed to complement vaccination and strengthen global efforts to control measles. The long-term objective of this research plan is to develop a safe and highly effective intranasal fusion protein (F) inhibitor as prophylaxis for use in high- risk unvaccinated populations. In preliminary work, we have generated potent MV fusion inhibitors by dimerizing peptides derived from the C-terminal heptad-repeat region of F and conjugating them to cholesterol. We have shown that prophylactic intranasal administration of our lead F fusion inhibitor efficiently protects suckling transgenic mice from developing fatal encephalitis after intranasal MV infection. By combining structure-based optimization of fusion inhibitors and modification of lipid components with virologic and in vivo assays, we aim to identify and characterize novel MV F fusion inhibitors that can be administered intranasally and have significantly improved anti-MV efficacy. To achieve our goals we propose two aims: 1. To use structure-guided mutagenesis and protein engineering to identify and develop optimized F peptide fusion inhibitors. A systematic mutational, biophysical and structural approach will identify and incorporate specific residue substitutions at the inhibitor-binding interface to impar additional binding energy. We will conduct in vitro and ex vivo studies to assess the antiviral activity of engineered F peptide fusion inhibitors. 2. To evaluate the protection afforded by optimized fusion inhibitors against MV infection in mice. We will evaluate the biodistribution and toxicity properties of the inhibitors, and use transgenic mouse models to assess their in vivo anti-MV potency. In an iterative process, the outcome of the experiments will guide further optimization, yielding a set of promising investigational anti-MV agents.